**4.1.1 Inoculum and wastewater**

Anaerobic granular sludge from a full-scale UASB reactor treated fruit juice processing wastewater in fruit juice industry, Olsztynek, Poland, was used as an inoculum for biohydrogen and methane production. Prior to inoculation of the hydrogenogenic reactor, the granular sludge was washed with three volumes of tap water and then boiled for 2 h to inactivate hydrogen consuming microorganisms. A final concentration of inoculum was 151 g TSS L-1. No pre-treatment of the granular sludge used for methane production was carried out prior to its inoculation in methanogenic reactor. Initial concentration of inoculum for methane production was 94 g TSS L-1. Both reactors were initially inoculated at a ratio of 20% (by volume).

UF whey permeate was obtained from a cheese production factory in Nowy Dwór Gdański, Poland. It was received from the factory once a week, was stored at -20°C and was thawed

Feasibility of Bioenergy Production from

d-1 and HRT of 3 d.

**4.1.4 Analytical methods** 

**4.2 Results and discussion** 

faded out.

measured with a water displacement meter.

Ultrafiltration Whey Permeate Using the UASB Reactors 213

the tank was maintained at 37°C by placing it in the thermostatic chamber. Overflow effluent flowed out in the top part of the storage tank and was collected in a separate container. The R2 was operated at an HRT of 3 d. The R2 performance (biogas production and composition in CH4, COD, TVFA concentration, pH) was monitored twice a week throughout the experimental period, from day 51 to 84. Before R2 was fed with R1 effluent, the diluted UF whey permeate had been used as a feedstock to reach the OLR of 2 g COD L-1

Determinations of COD, TSS, lactose, TVFA concentrations were carried out according to the Standard Methods for the Examination of Water and Wastewater (PN-74/C-04578.03; PN-78/C-04541; PN-67/A-86430; PN-75/C-04616.04). The measurement of the pH was continuously measured by the membrane electrodes, model ESAgP-301W, Eurosensor, placed in the liquid phase of reactor. Biogas composition (CH4, H2 and CO2) was analyzed by using an electronic analyzer (Gas Data GFM 430, Gas Data Ltd.). Biogas production was

The hydrogenogenic reactor (R1) operation started with a HRT of 24 h, OLR of 10 g COD L-1 d-1 and pH 5.8. Under these conditions hydrogen production was as low as 0.12 L H2 d-1 (0.027 L H2 L-1 d-1) (Fig. 13). It was noticed methane presence in biogas up to 19% v/v, while hydrogen concentration was still very low (up to 10% v/v). In order to inhibit methane production, the HRT was reduced to 12 h and OLR increased to 20 g COD L-1 d-1 on day 11. After 5 days, the HRT was increased to 24 h. Hydrogen content in biogas increased to the average value 15.7% v/v and methane was still present (<8% v/v). According to Yang et al. (2007), HRT shorter than 24 h does not favor the biohydrogen generation from cheese whey wastewater, but other researchers stated that short HRT could help to control the methanogenic reaction in hydrogenogenic phase (Castellό et al., 2009). Then, the HRT was again set at 12 h, OLR increased to 20 g COD L-1 d-1 and pH was decreased to 5.2 (day 24). It was seen that although the methanogenic bacteria was assumed to be washed out from the bioreactor under short HRT, the inhibition of methanogenic bacteria activity should be coupled with pH decrease. Liu et al. (2006) found that pH is the most critical factor for inhibition of methanogenesis and the optimum pH should be around 5.0 – 5.5. According to the literature, the optimum pH range for lactose (or whey) acidogenesis is between 6 and 6.5 (Venetsaneas et al. 2009). Antonopoulou et al. (2008) reported, that high concentration of hydrogen (over 25% v/v) in the gas phase was when the pH in the reactor was maintained at 5.2 ± 0.1. Wang et al. (2006) stated, that pH of 5.5 should be avoided in the biohydrogen fermentation process because at that level of pH, the propionic-acid type fermentation commonly occurred. The accumulation of propionic acid can lead to lower efficiency of methanogenic phase followed the hydrogenogenic phase. Mohan et al. (2007) found that pH 6 was the optimum for effective H2 yield. However, maintaining pH at 6 or above is difficult because of large amounts of fatty acids generation. At this study, the pH in the hydrogenogenic reactor was initially maintained on the level of 5.8. As the pH was reduced down to 5.2 (coupled with HRT shortening and OLR increasing), methane content in biogas

before used. The average composition of the feedstock was shown in Table 2. Prior to being fed into the reactor, the substrate was diluted with tap water to a COD concentration of 10,000 mg L-1 in accordance with the required OLR (10 or 20 g COD L-1 d-1) in the hydrogenogenic reactor. Diluting UF whey permeate was maintained at a temperature 4°C until used.
